NK cells are a subpopulation of lymphocytes whose unique receptors facilitate the detection of infected, transformed, or 'stressed' cells. This immune recognition subsequently leads to the development of NK cell- mediated cytotoxicity or the generation of cytokines and chemokines that activate other components of the immune system. Patients with hyper-IgE syndrome (HIES) have immune dysregulation and can be affected by recalcitrant cutaneous herpes virus and papillomavirus infections. A major genetic abnormality found in these patients is deletion or loss-of-function mutations in the gene encoding Dedicator of Cytokinesis 8 (DOCK8), a guanine nucleotide exchange factor (GEF) for Cdc42. The high incidence of recurrent cutaneous viral infections in DOCK8-deficient patients is suggestive of defects in natural killer (NK) cell function. In fact, others and we have recently shown that NK cells deficient in DOCK8 have reduced lytic function, decreased adhesion, F-actin accumulation at the cytotoxic synapse and an inability to polarize lytic granules toward the target cell. However, how DOCK8 can regulate so many critical steps in the development of NK cell killing is not known. It is our central hypothesis that the DOCK8-interactome coordinates the regulation of the actin and microtubule cytoskeletons to facilitate NK cell polarization and effect NK cellular cytotoxicity. Based on the preliminary data included in this proposal, we hypothesize that: (a) DOCK8 activation of CDC42 is critical to the development of NK cell killing; (b) DOCK8 interaction with WASP is critical for its localization and F-actin generation at the NKIS; (c) tali recruitment by DOCK8 mediates NK cell - target adhesion; (d) CCDC88B, a hematopoietically expressed protein that interacts with DOCK8 is involved in MTOC polarization and lytic granule clustering; (e) CCDC88B directly binds microtubules through its hook-like domain; (f) septins are DOCK8 interacting proteins that regulate NK cell killing through their effects on the microtubule and actin cytoskeletons. In order to test these hypotheses we will: (1) Determine the mechanism by which DOCK8 regulates F-actin dynamics and cell adhesion; (2) Define the role of CCDC88B during NK cell-mediated killing; (3) Determine the role of septins in the regulation of the NK cell cytotoxicity. The outcome of the proposed experiments will provide an experimental basis for understanding the molecular events that are involved in the regulation of NK cell effector functions, and will, in a broader context, advance our understanding of fundamental processes in cellular activation leading to F-actin regulation, MTOC polarization and granule exocytosis. Moreover, the information obtained through these studies will likely instruct how DOCK8 is functioning in other immune cells and provide new insight into its role in HIES.